Proteases are responsible, either directly or indirectly, for all bodily functions, including cell growth, differentiation and death (apoptosis), cell nutrition, intra- and extracellular protein turnover (housekeeping and repair), cell migration and invasion, and fertilization and implantation. These functions extend from the cellular level to the organ and organism level to produce cascade systems such as hemostatis and inflamation, and complex processes at all levels of physiology and pathophysiology.
Maintenance of vascular integrity is an important host response to injury. Complex hemostatis mechanisms of coagulation, platelet function, and fibrinolysis exist to minimize adverse consequences of vascular injury and to accelerate vascular repair. Many of these hemostatic mechanisms are initiated and/or regulated by cells of the wall of the blood vessel.
Tissue-factor-pathway inhibitor (TFPI) is a cell-surface associated glycoprotein which plays a key role in the regulation of tissue factor-initiated blood coagulation. Human TFPI is a trace 42-kDa plasma glycoprotein that is synthesized primarily by endothelial cells and consists of a negatively charged amino terminal region, three tandem Kunitz-type inhibitor domains, and a highly basic carboxyl-terminal tail (Wun, T. C., et al., J. Biol. Chem. 263:6001 (1988)). After a 22-residue signal peptide, the mature protein contains 213 amino acids with 18 cysteines. TFPI forms a complex with factor Xa and inhibits its amidolytic and proteolytic activity. The factor Xa-TFPI complex rapidly inhibits activity of the factor VIIa-tissue factor complex.
The cloning and characterization of a gene coding for a second tissue factor pathway inhibitor (TFPI-2), has been reported (Sprecher, C. A. et al., Proc. Natl. Acad, Sci. USA, 91, 3353-3357 (1994)). This gene was initially identified on the basis of primary sequence homology and structural similarity. Subsequent characterization has confirmed its predicted activity as a protease inhibitor. Alterations of the hemostatic system can result from such causes as neoplasia and trauma. Such alterations result in an increased incidence of thrombotic disorders such as venous thrombosis, pulmonary embolism, atrial fibrillation, cerebral thrombosis, and hemophilia. Thus, there is a need for identification and characterization of polypeptides that function as inhibitors of the coagulation pathway which can play a role in detecting, preventing, ameliorating or correcting such disorders.